A limited number of proteins have been identified as being primarily associated with bone tissue. In addition, reports in the literature have presented compelling evidence that the targets of these proteins are receptors on the cell surface. Accordingly, applicants anticipate that protein domains may be responsible for targeting these proteins to cell receptors or receptor ligands in the extracellular matrix of the target tissue. Therefore these peptides could be used for a wide variety of therapeutic uses including the delivery of drugs that could help in the treatment of musculoskeltal disorders, genetic or acquired, osteoporosis and metastatic cancer. Furthermore it is anticipated that the peptides themselves can serve as therapeutic agents providing osteogenic or trophic activity for osteoblast, mesenchymal or hematopoietic cell lineages.
To detect peptides that are targeted to the bone, a phage display peptide library was prepared using a kit from New England Biolabs. The phage display library provides a selection technique wherein a peptide is expressed as a fusion protein with a bacteriophage coat protein, resulting in the display of the fused protein on the surface of the virion, while the DNA encoding that fusion protein resides within the virion. The phage peptide library is created using random nucleic acid sequences to generate fusion proteins that comprise the coat protein linked to a random peptide sequences. An in vitro or in vivo selection process designated “biopanning” is then conducted to identify those randomly generated peptides that bind to the target tissue. More particularly, the phage display library is contacted with the target cells, the unbound phage are washed away and the specifically bound phage are eluted. The eluted phage is then amplified and taken through addition binding/amplification cycles to enrich the pool in favor of binding sequences. Applicants have identified 14 peptides that are bone targeting peptides (see Example 1) and these peptides form the basis of the present invention.